Degradation and remodeling of the ECM are essential processes for normal repair after tissue trauma. The physiological response to tissue trauma is a complex process involving multiple factors including cell migration and replication, turnover of extracellular matrix (ECM) components and changes to the cellular microenvironment. Essentially, such a response involves the repair or replacement of damaged tissues. The precise nature of such repair or replacement depends upon the tissues involved, although all such processes involve certain basic principles. The normal and necessary repair of any tissue after any trauma requires the coordination of a wide array of factors by regulated gene expression.
Fibrosis is therefore typically a reaction to tissue trauma. A number of different factors are believed to affect or modulate the biological pathways or mechanisms leading to tissue fibrosis. Such factors may include early inflammatory actions, a local increase in fibroblast cell populations, modulation of the synthetic function of fibroblasts, and altered regulation of the biosynthesis and degradation of collagen.
The pathophysiological response to the tissue trauma seen in fibrosis results in the formation of abnormal tissues which do not duplicate the functionality of the original organ tissue, so that the repair of tissue trauma does not lead to a complete restoration of organ capacity and function. One example of a fibrotic process which results from pathophysiological responses to tissue trauma is cardiac fibrosis. Cardiac fibrosis has a number of causes, which lead to the deposition of fibrotic tissue. For example, cardiac fibrosis may result from heart failure, hypertension and other cardiac pathological/disease states. As the deposition of such fibrotic tissue increases, the ability of the heart to function decreases, leading to disability and eventually death of the patient. The formation of fibrotic tissue in the heart is characterized by the deposition of abnormally large amounts of extracellular matrix components, including collagen, as well as other matrix proteins. Therefore, the cardiac fibrotic process needs to be inhibited in order to prevent damage to the cardiac tissue and hence to the ability of the heart to function.
Cardiac fibroblasts are important to the cardiac fibrotic process because they produce interstitial proteins and other myocardial components which have been implicated in heart failure (Hess et al, Circ., 63:360-371 (1981); Villari et al, Am J. Cardiol., 69:927-934 (1992); Villari et al, JACC, 22:1477-1484 (1993); Brilla et al, Circ. Res., 69:107-115 (1991); and Sabbah et al, Mol. & Cell Biochem., 147:29-34 (1995)).
The pathology of heart failure is clearly associated with fibrosis for a number of cardiac pathological or disease states, including those associated with both volume and pressure overload (Maron et al, Am. J. Cardiol., 35:725-739 (1975); Schwarz et al, Am. J. Cardiol., 42:661-669 (1978); Fuster et al, Circ., 55:504-508 (1976); Bartosova et al, J. Physiol., 200:285-295 (1969); Weber et al, Circ., 83:1849-1865 (1991); Schaper et al, Basic Res. Cardiol., 87:S1303-S1309 (1992); Boluyt et al, Circ. Res., 75:23-32 (1994); and Bishop et al, J. Mol. Cell Cardiol., 22:1157-1165 (1990)). Cardiac surgery also may cause cardiac fibrosis; such fibrosis may also lead to the requirement for an additional operation, which is often associated with higher morbidity and mortality.
Detection and/or quantitation of cardiac fibrosis is therefore very important for preventing or treating such fibrosis. Various imaging techniques may be used to see the effects of cardiac fibrosis, but actually detecting such fibrosis at the molecular level currently requires an invasive procedure to obtain a tissue sample (biopsy), as there are no commercially available non-invasive tests for detection of cardiac fibrosis at the molecular level.
BNP (Brain Natriuretic Peptide) belongs to a family of natriuretic peptides produced by the heart. BNP and its related natriuretic peptide ANP contain a 17-amino acid ring structure and are produced by the cardiac atria in response to volume overload and by ventricles in response to pressure overload, respectively (Broomsma et al., 2001. Cardiovascular Research 51, 442-449.; McCullough et al., 2003. Reviews in cardiovascular medicine 4, suppl. 7, S3-S12). These hormones have powerful diuretic, natriuretic, vascular smooth muscle relaxing and vasodilation actions, thus lowering blood volume and blood pressure (Azzay et al., 2003. Heart Failure Review 8, 315-320.). With its impressive physiologic actions, BNP was an attractive target for development as a therapeutic agent for heart failure and/or as a diagnostic marker.
Both ANP and BNP are formed as pre-pro-polypeptides. Human BNP is derived from the 134-aa precursor preproBNP. Upon stimulation of release, a 26-aa signal peptide sequence is cleaved from the N-terminus of preproBNP. During release into circulation, the remaining proBNP1-108 prohormone is further cleaved by corin, a membrane-bound serine protease, into an N-terminal pro-BNP1-76 fragment and the active 32-peptide, C-terminal proBNP77-108 hormone termed BNP (Azzay et al., 2003. Heart Failure Review 8, 315-320.). The principal function of ANP and BNP is to protect the cardiovascular system from volume overload. They are secreted in response to the wall stretch, ventricular dilation and/or increased pressures resulting from fluid overload (Azzay et al., 2003. Heart Failure Review 8, 315-320.). Both cause intravascular volume contraction by inducing a shift of fluid from the capillary bed to the interstitium, resulting in a decrease in preload and blood pressure. BNP has the important ability to decrease left ventricular filling pressures without a resultant reflex tachycardia, reflex vasoconstriction, and further activation of vasoconstricting neurohumoral systems. BNP also has lusiotropic effects and has been demonstrated to inhibit cardiac fibrosis. The natriuretic peptides also appear to exhibit an antimitogenic effect in the heart and other organ systems, suggesting a potential role in the modulation of cell growth. Additional evidence suggests a direct vasodilatory effect on the coronary arteries with a reduction in myocardial oxygen consumption.
In addition to these cardiac and vascular properties, BNP has a direct effect on renal hemodynamics and function. Increased glomerular filtration is the result of an unbalanced vasodilatation of the afferent arterioles and vasoconstriction of the efferent arterioles. There also appears to be a direct tubular effect on sodium and water handling, resulting in natriuresis and diuresis as well as inhibition of aldosterone and rennin release. The net effect of these properties is balanced vasodilatation of the arterial and venous beds as well as natriuresis and diuresis (Fonarow G. C, 2003. Heart Failure Review 8, 321-325.).
Most effects of ANP and BNP are mediated through binding to the A-type natriuretic peptide receptor, which activates guanyl cyclase, leading to the formation of cyclic guanosine monophosphate (cGMP). CGMP has potent vasodilatory actions and acts as a second messenger for BNP. The A-type natriuretic peptide receptor is expressed in a variety of tissues, including kidney, blood vessels, adrenal glands, heart, lungs, adipose tissue, eye, pregnant uterus and placenta. Clearance of ANP and BNP from the blood is effected in two ways: through a special clearance receptor, the C-type natriuretic receptor, and through enzymatic degradation by neutral endopeptidases (Broomsma et al., 2001. Cardiovascular Research 51, 442-449). The natriuretic peptides are characterized by a 17 aa central ring structure, which is formed by a disulfide bridge and is suggested to be necessary for the binding of these peptides to their respective receptors and for their biological activity (Azzay et al., 2003. Heart Failure Review 8, 315-320.).
ANP and BNP are both expressed more in Atria than Ventrcles. BNP has a more favorable expression in ventricles (atria:ventricle expression ration 3:1 compared with 40:1 to ANP). In a situation of a failing heart both BNP and ANP expression is increased 100-fold above normal levels (Trends Endocrinol Metab. 2003 Nov;14(9):411-6). However, BNP rise is often larger and more rapid than ANP, and it emerged as a superior marker for heart failure and left-ventricular dysfunction (Azzay et al., 2003. Heart Failure Review 8, 315-320.). BNP was shown to have a diagnostic benefit for few different clinical purposes: 1. Elevated plasma levels of BNP are found in conditions of increased cardiac wall stress. In congestive heart failure (CHF), circulating concentrations of BNP are clearly elevated and this elevation reflects the severity of the condition. Therefore it can be used to establish prognosis in patients with heart failure. In addition, it provides a tool to monitor changes in the severity of failure without using more sophisticated diagnostic modalities like Echo imaging.
2. Several studies have clearly shown that natriuretic peptides are excellent prognostic indicators for survival in heart failure.
3. Detection of response to treatment: a continued increase in plasma concentration would be indicative of unsuccessful, and a decrease of successful, treatment. 4. In myocardial infarction ANP and BNP concentrations are also elevated, and are of prognostic value for indicating patients most at risk. 5. BNP measurements help to differentiate between cardiac versus non-cardiac causes of dyspnea (Lancet. 1994 Feb. 19;343(8895):440-4). However there is limited evidence that it can be a useful marker for the very early detection of cardiac damage or heart failure when patients are still asymptomatic.
The N-terminal proBNP (ntBNP) is more stable and its serum levels rise more then BNP, therefore it is theoretically a better marker for BNP overproduction than BNP itself. Yet, its added value over BNP diagnostic wise is minor. When studying CHF population with left ventricular ejection fraction (LVEF)<40% BNP and ntBNP showed no difference in the diagnostic properties. For patients with LVEF<50% (less severe patients), ntBNP showed slight improvement over BNP—receiver operating characteristics area under curve of 0.82 for ntBNP and 0.794 for BNP (Eur J Heart Fail. 2004 Mar. 15;6(3):295-300).